Aplastic anemia (AA) and other types of bone marrow failure have clinical and laboratory features consistent with an autoimmune pathophysiology, with a diversity of inciting antigens, including viruses, chemicals, and drugs. Whatever its specific etiology, a majority of patients respond with hematologic improvement after immunosuppressive therapies. One important clinical feature of AA is its evolution, sometimes years after normalization of blood counts, to other hematologic diseases such as paroxysmal nocturnal hemoglobinuria (PNH), which derive from clones of hematopoietic stem cells. Our laboratory studies have focused on the immune pathophysiology of AA, identification of a viral antigen, and the mechanism of late clonal evolution. Studies of etiology have continued to focus on an unknown hepatitis virus in the post-hepatitis AA syndrome (see Z01 HL 02319-14 HB). Current studies of the immune systems role in bone marrow suppression have focused on gamma-interferon (gamma-IFN), a lymphokine that inhibits hematopoiesis in vitro and in vivo. In our animal model for immune-mediated AA, congenic lymphocytes induce profound marrow aplasia, which can be abrogated by early administration of anti- lymphocyte globulin (ATG), cyclosporine, and monoclonal antibody to gamma-IFN. Current efforts are directed to producing chronic aplasia by periodic infusions of lymphoid cells, as well as determination of the specificity of the immune response. We have measured gamma-IFN in circulating and marrow lymphocytes using flow cytometry. About 2/3 of patients with severe AA and also with the diagnosis of hypocellular myelodysplasia shown the presence of this cytokine in blood T cells; less than 10% of recovered patients scored positive. The presence of the cytokine was highly predictive for response to immunosuppressive therapies: 16/16 patients whose cells contained gamma-IFN responded, compared to 4/12 patients whose blood lacked gamma-IFN; IFN declined after treatment, while IL-4 content increased. Recurrence of IFN intracellularly predicted relapse. Marrow intracellular IFN may be even better correlated with response. Measurement of surface gamma-IFN allows isolation of viable activated T cells, which may be useful in determining lymphoid clonality and characterizing and even identifying their antigens. Intracellular cytokine measurements indicating TH1/TH2 balance are also altered by in vitro treatment with cyclosporine, various androgen preparations, and growth factor combinations; stem cell factor and granulocyte colony stimulating factor also depress IFN expression in vivo in normal individuals undergoing stem cell mobilization. In other clinical studies in aplastic anemia, 24 patients with newly diagnosed severe aplastic anemia have now been randomized in a comparative protocol; the response rate has been higher for ATG than for high dose cyclophosphamide, but frank relapses and cytogenetic abnormalities have occurred only in the ATG arm. For children, our standard protocol has been modified to shorten the duration of cyclosporine therapy, begin cyclosporine is begun later in order to avoid abrogation of ATG tolerization, and the new immunosuppressive agent mycophenolate mofetil has been added, all in an effort to decrease the high relapse rate. In the laboratory, late clonal disease, both PNH and myelodysplasia, using sensitive flow cytometric and fluorescent in situ hybridization assays. Approximately 20% of patients with AA and probably a larger proportion of cases of hypocellular myelodysplasia present with evidence of a PNH clone. Our results are consistent with the hypothesis that PNH represents an escape mechanism in immune-mediated bone marrow failure, and that two steps are required for the development of PNH. Alternatively, a glycosylphosphoinositol- linked protein may be involved in the initial antigenic stimulation. - autoimmunity, T cells, bone marrow failure, myelodsyplasia, immunosuppression, interferon - Human Subjects